Figure 6. Dynamin oligomerization is dependent on direct actin-dynami interactions and the status of the actin cytoskeleton.

a) Cos-7 cells were infected with viruses expressing Dyn1^K/E, Dyn1^E/K or Dyn1^WT. When indicated, cells expressing Dyn1^WT were treated with 0.2 µM Lat A or 1 µM Cyto D for 20 min prior to fixing. 24 hours after infection cells were fixed and stained using monoclonal anti-GTPase antibody conjugated with Alexa Fluor 488 (donor fluorophore) and monoclonal anti-PRD antibody conjugated with Alexa Fluor 594 (acceptor fluorophore). Grey images represent intensity of Alexa Fluor 488 and are overlaid by positive FRET.

Lifetimes are shown as discrete colors. Fast lifetimes between 0–2200 ps are shown in red (positive FRET) and slow lifetimes above 2200 ps are shown in blue (no FRET).

b) Histogram of fluorescence lifetimes (τ1) of the donor fluorophore (Alexa Fluor 488) in the whole image after the lifetimes were curve fitted to a two exponential decay curves with one exponential fixed at the average lifetime for donor only fluorophore (τ2=2700 ps).